The chlorooxide (ClOx-) ions are anthropogenic biocides, bleaches, and oxidants that, because of their high solubility and ubiquitous use by agriculture, industry, and the military, have become major contaminants of drinking water and fresh water environments. The various chlorooxides are both suspected and documented health hazards. The following proposal will determine the feasibility of in-depth studies of microbe-based ClOx- detoxification at the protein/enzyme level. The U.S. EPA has included chlorite (CIO2-) on its list of Primary Drinking Water Standards, with a legally mandated upper limit of 0.8 mg/L in drinking water. Health risks associated with chronic CIO2- exposure include anemia and nervous system disorders in children and infants. After its recent identification as a well water contaminant, perchlorate (CIO4-) was added to the EPA's Contaminant Candidate List. Its full health effects are currently under study by the EPA and by others. In response to an influx of chlorooxides, nature has evolved mechanisms to detoxify and even exploit them. Microorganisms that anaerobically respire chlorooxides, including CIO4-, CIO3-, and CIO2-, have been isolated over the last decade. Traditional means of removing small ions from water are expensive or nonspecific, so bioremediation of chlorooxides is an emerging bioengineering goal. Dechloromonas aromatica, the only (ClOx-)-respiring microbe with a sequenced genome, is the obvious choice for biochemical study (J.D. Coates, University of California, Berkeley). Moreover, Dechloromonas is unique among strains in pure culture, in that it oxidizes intransigent hydrocarbons including benzene, a major environmental contaminant and a potent human carcinogen. We seek [1] to isolate, partially sequence, and identify the genes encoding (ClOx-)-metabolizing activities in D. aromatica; [2] to obtain sufficient quantities of pure enzymes for mechanistic and X-ray structural studies from the host and heterologous expression systems; and [3] to partially characterize the endogenous and expressed enzymes. These objectives set the stage for future mechanistic, spectroscopic, structural, and protein-engineering studies of these components of the (ClOx-) detoxification pathway. This work will be part of a more global effort aimed at developing bioremediation systems, and/or enzyme-based sensors for environmental monitoring.
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